Bacterial Fc receptors have been identified by their ability to bind to a site within the constant region of various classes and subclasses of mammalian IgG (Myhre, E.B. and G. Kronvall. 1981. Immunoglobulin specificities of defined types of streptococcal Ig receptors. In: Basic Concepts of Streptococci and Streptococcal Diseases [Holm, S.E. and P. Christensen, eds.]pp. 209-210, Reed Book, Ltd., Chertsey, Surrey) The Fc region of the IgG antibody molecule is associated with the biological effector properties of the molecule while the antigenic recognition elements are located in the two identical Fab portions of the antibody. Consequently, the interaction of bacterial Fc receptors with constant region determinants on the heavy chain of IgG does not interfere with the ability of the antibody to recognize its antigen; it is this property that makes these receptors so useful as tracers of antibody-antigen interaction (See Boyle, M.D.P. (1984) "Applications of bacterial Fc receptors in immunotechnology," Biotechniques 2:334-340.)
To date, six types of bacterial receptors have been described based on the reactivity of whole bacteria with different classes and subclasses of mammalian immunoglobulins. The most extensively characterized receptor is the type I receptor isolated from Staphylococcus aureus, and more commonly designated protein A. The type II receptor is found on a few group A streptococci. The type III receptor, also known as protein G, is present on many group C streptococci and on some human group G streptococci The type IV receptor is isolatable from a few bovine group G streptococci, the type V receptor is found on certain Streptococcus zooepidemicus strains, and the type VI receptor has been recently reported on selected strains of S. zooepidemicus
The efficient recovery and purification of bacterial receptors from wild-type bacteria, i.e., non-recombinant bacteria, are critical to the ultimate commercial use of these receptors. The best prior art process known for the recovery of protein G from group G or group C streptococci uses a proteolytic enzyme extraction procedure. See Reis, K. J., Ayoub, E. M., and Boyle, M. D. P. (1985) "A rapid method for the isolation and characterization of a homogenous population of streptococcal Fc receptors," J. Microbio. Methods 4:45-58; Bjorck, L. and Kronvall, G. (1984) "Purification and some properties of streptococcal protein G, a novel IgG-binding reagent," J. Immunol. 133:969-974; and European Patent Application No. 0 131 142, published on Jan. 16, 1985. The invention concerns the use of proteolytic enzymes, e.g., papain, trypsin and pepsin, to solubilize the protein G bound to the streptococcal cell wall. The EPO application also discloses U.S. Pat. No. 3,850,798 which uses trypsin to recover protein A.
Wild type protein G is distinguishable from the recombinant form of the protein by having the ability to bind not only to the Fc region of human IgG, but also to human albumin.
The prior art processes all use enzymatic hydrolysis as the method to extract either protein A or protein G. We have tried the trypsin extraction process to recover protein G from wild-type group G or group C streptococci and have been able to obtain a yield of ca. 10 mcg. of protein G from a gram of wet bacterial mass. Though this is an acceptable product yield, there is still a need to have a more efficient extraction process. Further, the proteolytic enzymes may degrade the protein G product. Thus, the prior art methods of extracting protein G from wild-type streptococcus may produce low molecular weight degraded forms of protein G, which are not as stable and cannot be radiolabeled without loss of functional activity (see Boyle, Michael D. P. and Reis, K. J. [1987]"Bacterial Fc receptors," Biotechnology 5:697-703).